Development of a genotyping protocol for Mojave desert tortoise scat

Noninvasive fecal genotyping can be a useful tool for population monitoring of elusive species. We tested extraction protocols on scat samples from the threatened Mojave Desert tortoise, Gopherus agassizii, to evaluate whether scat-based mark–recapture and population genetic monitoring studies are feasible.We extracted DNA from G. agassizii scat samples collected in California and Nevada using several extraction protocols and evaluated the reliability of resulting genotypes using quality scores, maximum likelihood reliability estimates, and paired scat and blood genotypes from the same individuals. Finally, we assessed probabilities of identity and sibship, and locus amplification quality, and calculated genotyping error rates for 19 microsatellite loci to determine the best set of loci to use with G. agassizii scat extractions. We found that genotype quality depended more on the sample quality than on the extraction method, and that the Qiagen DNeasy Plant Mini extraction kit is an efficient method for extracting tortoise DNA from tortoise scat. We identified 6 G. agassizii microsatellite loci that can be used to generate a unique molecular tag for individual tortoises. We characterized the reliability of an additional 13 microsatellite loci for use in population genetic analyses where additional power at the expense of some increase in error may be advantageous. As proof of concept, with very low error rates, we matched 3 opportunistically collected scat samples to blood genotypes from animals captured during population surveys within the study area and discovered at least 3 new individuals, even after 2 yrs of extensive survey work. These results suggest that genotyping of field-collected scat can complement existing methods used in long-term demographic and movement studies of G. agassizii and other, closely related, tortoise species.